Effects Of Astaxanthin On The Expression Of Pyroptosis-Related Proteins After Intestinal Ischemia-Reperfusion In Mice

Objective:Evaluate the effect of astaxanthin（AST）on intestinal ischemia-reperfusion（II/R）injury and reveal the potential mechanisms of astaxanthin in Balb/c mice.Methods: Twenty-four healthy male balb/c mice aged 7-10 weeks were randomly divided into 3 groups（n=8）by random number table method: sham operation（Sham）group,ischemia-reperfusion（I/R）group,andastaxanthin（AST）group.Seven days before modeling,the AST group was given astaxanthin 30 mg/kg/d by gavage at 9 am every day,and the Sham and the I/R groups were given 10 ml/kg /d olive oil by gavage.On the seventh day,the I/R group and the AST group were used to establish the model of intestinal ischemia-reperfusion injury by clipping the superior mesenteric artery（SMA）for 45 minutes and reperfusion for 2 hoursafter gavage40 minutes.The Sham group was used as a normal control group,only the superior mesenteric artery was exposed without clipping.After successful modeling,the mouse orbital vein blood was collected and the level of interleukin-1β（IL-1β）and IL-18 were detected by enzyme-linked immunosorbent assay（ELISA）.After being sacrificed,5 cm of small intestinal tissue was taken from the ileocecal valve to the heart,the pathological changes of intestinal mucosa were observed by HE stainingand Chiu’s score.The content of malondialdehyde（MDA）,glutathione peroxidase（GSH-Px）and superoxide dismutase activities（SOD）in intestinal tissue were determined by ELISA.Western Blot was used to detect the expression of nucleotide-binding oligomerization domain-like receptor protein 3（NLRP3）,caspase1 in intestinal tissue（caspase-1）and gasdermin-D（GSDMD）protein in intestinal tissue.Results: 1.Compared with the Sham group,the levels of serum inflammatory factors IL-1β and IL-18 in the I/R group were significantly increased（p < 0.05）;the intestinal mucosa was significantly damaged,the Chiu,s score was significantly increased,and the pathological characteristicsbetween the groups was different（p<0.05）.The content of MDA in intestinal tissue was significantly increased,the expressions of NLRP3,caspase-1 and GSDMD proteins were notably up-regulated,while the activities of GSH-Px and SOD were decreased（p< 0.05）.2.Compared with the Sham group,the levels of serum inflammatory factors IL-1βand IL-18 in the AST group were increased（p<0.05）,and the intestinal mucosal injury was mild and the Chiu’s score was increased（p<0.05）.Similarly,the content of MDA in the intestinal tissue increased,the activities of GSH-Px and SOD decreased（p<0.05）.Besides,there was no significant differenceof NLRP3,caspase-1 and GSDMD in protein expression.3.Compared with the I/R group,the levels of serum pro-inflammatory factors IL-1β and IL-18 in the AST group were significantly lower,with an average value of130.2 pg/m L and 99.12 pg/m L respectively,and there was a significant difference between the two groups（p< 0.05）.The degree of intestinal mucosal injury was significantly reduced and the Chiu’s score was also significantly reduced（p<0.05）.The content of MDA in the intestinal tissue was significantly decreased,but the activities of GSH-Px and SOD were notably increased（p< 0.05）.Western blot showed that the expressions of NLRP3,caspase-1 and GSDMD proteins were significantly down-regulated（p< 0.05）.Conclusions:Astaxanthin pretreatment can alleviate intestinal I/R injury in mice,and the mechanism may be related to inhibiting oxidative stress and reducing the expression of pyroptosis-related proteins.